Proteins move in the nucleus and transiently interact with binding sites there, but in most cases we do not know why they are so mobile or what they are bound to. Our work has focused on using FRAP to investigate the mobility of transcription factors both at specific promoter sites and also at other generic sites throughout the nucleus. In much of this work, we have used a mouse cell line containing a GFP-tagged glucocorticoid receptor (GFP-GR), which we were able to visualize in live cells binding to a tandem array of MMTV promoter target sites. FRAP of GFP-GR at this site revealed rapid exchange of GFP-GR with full fluorescence recovery in less than a minute, even though transcription persists for several hours. This surprising result raised questions about both the mechanism and purpose of rapid exchange. Our recent work has begun to address these questions by showing that chaperones and proteasomes are involved in regulating exchange, and that exchange rate appears coupled with transcription. In related studies in yeast, we have found that transcription factor mobility throughout the nucleus reflects non-specific DNA binding and these dynamic interactions are accelerated by a chromatin remodeler. To better understand FRAP recovery data and extract quantitative information about binding from these data, we are developing mathematical models to account for FRAP recoveries in the presence of diffusion and binding interactions.
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